Combined inhibition of polyamine biosynthesis and uptake blocks Candida albicans virulence

Polyamines, small organic polycations, are essential for life, and cells maintain polyamines via synthesis and uptake. Recently, we identified Hol1 as the high-affinity polyamine transporter in Saccharomyces cerevisiae. Hol1 is a conserved fungal-specific transporter with Candida albicans having two HOL1 homologs (orf19.4889 and orf19.2991) and no identifiable HOL1 homolog in mammals. Deleting both HOL1 homologs blocked efficient polyamine uptake in C. albicans, establishing Hol1 as the high-affinity polyamine transporter in C. albicans. Combined deletion of HOL1 and SPE1, encoding ornithine decarboxylase (ODC), the first enzyme in the polyamine synthesis pathway, resulted in a severe growth defect, confirming the importance of polyamines for C. albicans growth. In addition, cells lacking HOL1 and SPE1 failed to form hyphae when exposed to serum, suggesting a role for polyamines in C. albicans virulence. In accord with this hypothesis, in the mouse model of disseminated candidiasis, the homozygous spe1Δ mutant, like the WT strain, readily colonized the kidney and all mice died within 2 weeks following intravenous inoculation. In contrast, the homozygous spe1Δ hol1Δ mutant was avirulent with all mice surviving the infection. Consistent with these genetic results, simultaneously treating WT cells with l-α-difluoromethylornithine, an irreversible inhibitor of ODC, to inhibit polyamine biosynthesis and with the polyamine transport inhibitor Trimer44NMe to inhibit uptake substantially impaired C. albicans growth and hyphal differentiation. We conclude that polyamines are critical for C. albicans virulence and could be of potential therapeutic interest via combined targeting of polyamine synthesis and the fungal-specific polyamine transporter Hol1.IMPORTANCEFungal infections are a growing concern, and a predominant human opportunistic pathogen is the fungus Candida albicans. Current antifungals commonly target cell wall and cell membrane biosynthesis or integrity; however, resistant strains are emerging. Polyamines are essential small organic cations that cells synthesize and import. We identify polyamines as a possible new target for antifungal therapies. The Hol1 polyamine transporter is unique to fungi and is distinct from mammalian transporters, so it is an intriguing antifungal target. As proof of concept, we show that combined knock-out of Hol1 and a polyamine biosynthesis gene impairs C. albicans growth and hyphal differentiation in culture and virulence in mouse infection assays. Moreover, we identify a polyamine analog that robustly inhibits Hol1 function, providing insights into potential new therapeutics.